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1. Field of the Invention
The present invention relates to a method for removing or blocking toxic carbonyl containing compounds and/or dicarbonyl containing compounds to prevent them from otherwise binding to and/or cross-linking proteins, forming protein adducts and/or cross-linked complexes, denaturing proteins, disrupting protein structure and/or function, and/or from causing disease states produced by or associated with carbonyl- and/or dicarbonyl-induced protein complexes. As is shown herein, the present invention is also applicable when applied in vitro, for example, to bovine serum albumin, but not limited to that usage. More particularly, the present invention relates to a method of using D- and/or L-arginine, or substituted or modified arginine, or arginine-containing compounds to remove or block toxic carbonyls and/or dicarbonyls, for example in vivo. Still more particularly, the present invention relates to a method of treating disease states which are associated with levels of toxic carbonyls and/or dicarbonyls in vivo, such as diabetes mellitus and acute chemical poisoning, by removing or blocking toxic carbonyls and/or dicarbonyls with the therapeutic administration of L- and/or D-arginine; for example, one embodiment of the present invention relates to the treatment of diabetes mellitus by reducing the in vivo level of toxic dicarbonyl-containing methylgloxal and related toxic metabolites of sugar in a patient by administering L- and/or D-arginine to a patient.
The present invention relates also to a method for removing, blocking, or scavenging dicarbonyls and carbonyls in vitro. More particularly, the present invention relates to a method of using D-arginine and/or L-arginine, or arginine-containing compounds to remove (scavenge or attach to) or block carbonyls or dicarbonyls.
2. General Background of the Invention
Reactive carbonyl groups, for example in vivo are toxic by, inter alia, reacting with native proteins to form adducts and/or cross-linked complexes. This process can inactivate important proteins as well as form unwanted protein complexes in vivo. Indeed, a number of ailments are believed to be caused by the accumulation in vivo, and subsequent reaction of toxic carbonyls and/or dicarbonyls and toxic carbonyls and/or dicarbonyl-containing compounds with native compounds such as proteins. For example, it, is believed that at least some complications associated with diabetes mellitus, such as, for example, cataracts and kidney problems, are related to the accumulation in vivo, and reaction of toxic dicarbonyl-containing compounds that are sugar-derived, such as, for example, but not limited to, methylglyoxal, glyoxal, deoxyglucosone and chemicals of similar structure. It is believed that, for example, in diabetes mellitus, high blood glucose levels can lead to high levels of methylglyoxal. The methylglyoxal can then, via its reactive dicarbonyl group, react with native proteins leading to, inter alia, unwanted protein-methylglyoxal adducts and cross-linked proteinaceous complexes. These complexes can then be responsible for such symptoms of diabetes mellitus as cataracts and kidney problems.
The ocular structures of higher vertebrates vary structurally and chemically from those of humans. For example, the human eye lacks the tapetum lucidum of many higher vertebrates, for example the deer. The human sclera contains no bones as in birds. Human aqueous humor does not coagulate as in the rabbit. The human retina contains color receptor pigments lacking in the dog. Prior to the publication of the inventor""s exhaustive biochemical and enzymatic study of the human eye no one had isolated methylglyoxal, glyoxalase I and glyoxalase II from the human lens (Haik et al. 1994, which is incorporated herein by reference).
Whether diabetic or not the human lens is never vascularized and is completely dependent on anaerobic glycolysis. Accordingly, both diabetics and non-diabetics produce levels of methylglyoxal in the human lens many times higher than normal blood levels. Our experiments with bovine serum albumin have shown that methylglyoxal has the ability to produce solid yellow gel formation from liquid proteins. The inventor attributes this at least primarily, and not intending to be bound by theory, to imine bond formation and the cross-linking of proteins.
As discussed in more detail below, the inventor has discovered that one can block methylglyoxal-induced gel formation of liquid bovine serum albumin by pretreatment with D-arginine or L-arginine in the free base or hydrochloride form. The inventor believes, but does not intend to be limited by any particular theory, that a process involving protein cross-linking by methylglyoxal (and perhaps to a lesser degree by glyoxal and other dicarbonyls) can produce rapid cataract formation in diabetics and relatively slower senile cataracts formation in non-diabetics over a period of years. Larger amounts of methylglyoxal would be expected to be produced from higher concentrations of the substrate glucose, however, free methylglyoxal levels are rapidly diminished by attachment to available proteins and amino acids or detoxified via the glutathione-dependent glyoxalase system. This is especially important since levels of reduced glutathione diminish with oxidative stress and the aging of tissues.
Increased levels of toxic carbonyl-containing compounds associated with high glucose levels, such as methylglyoxal, can also form adducts in the blood which can lead to kidney problems. Additionally, occupational or accidental exposure to toxic carbonyl-containing compounds can cause any number of medical complications associated with the formation of protein adducts within the body such as, for example, cataracts, arthritis, kidney, lung and circulation problems (not to exclude circulation problems in the retina referred to as diabetic retinopathy, and not to exclude vasculopathy elsewhere in the extremities) and so forth. Finally, it is believed that at least some of the physiological changes associated with aging, such as senile cataracts, are related to adduct formation caused by such toxic agents as toxic dicarbonyls.
It is therefore desirous to devise a method of removing and/or blocking toxic carbonyls and/or dicarbonyls from, for example in vivo environments before they react with native tissues to form adducts and/or detrimental cross-linked complexes. However, prior to the present invention, such a method was not known, making adduct formation and cross-linking formation from toxic carbonyls and/or dicarbonyls a problematic clinical and/or aging phenomenon.
The present invention provides a method for removing toxic carbonyls and/or dicarbonyls from environments, for example in vivo environments, before they react with tissues to form adducts and/or detrimental cross-linked complexes, thereby providing a method for eliminating or reducing the detrimental effects caused, for example in vivo, by toxic dicarbonyls.
The present invention recognizes that both L- and D-arginine are reactive with toxic carbonyls and dicarbonyls in such a manner that the presence of D- and/or L-arginine can react with toxic carbonyls and dicarbonyls in order to block and/or remove them before they can react with other compounds, such as native proteins. The present invention further recognizes that L- and/or D-arginine can, for example in vivo, effectively compete with native xe2x80x9ctargetxe2x80x9d compounds, such as proteins, for binding to any toxic dicarbonyls and carbonyls that might be present, thereby providing a method for blocking and/or removing toxic carbonyls and/or dicarbonyls from an environment before the dicarbonyls and/or carbonyls can react with native tissues and cause damage.
The present invention further recognizes that in addition to physiologically active L-arginine, importantly one can also use non-naturally occurring D-arginine with the instant invention. The use of D-arginine to block and/or remove toxic dicarbonyls and/or carbonyls from, for example in vivo environments provides a non-physiologically reactive substance with which to block and/or remove toxic dicarbonyls and/or carbonyls. This can be of great value as it provides a means of administering a water soluble and excretable scavenger that is not physiologically active other than to act in the blocking/scavenging manner of the present invention.
The method of the present invention solves the problems confronted in the art in a simple and straightforward manner. The present invention recognizes that D- and L-arginine can, for example in vivo, reduce the level of toxic carbonyls and/or dicarbonyls and thereby reduce or prevent adduct formation and cross-linking with native tissues which would otherwise be caused by the presence of toxic dicarbonyls and/or carbonyls in a living body. What is provided therefore is a method which utilizes arginine, and/or substituted or modified arginine, to preferentially and chemically react with toxic carbonyls and dicarbonyls, preferably in vivo, to thereby remove them before they react with native tissues to form detrimental adducts and/or cross-linked complexes. This method can reduce the level, and/or block toxic carbonyls and dicarbonyls in a living body and thereby reduce the damaging effects caused by cross-linking and/or adduct formation of carbonyls with native tissues.
It is further recognized and an aspect of the present invention that in addition to physiologically active L-arginine, importantly one can also use non-naturally occurring D-arginine to block and/or remove toxic dicarbonyls and/or carbonyls from, for example in vivo environments. The use of D-arginine importantly provides a non-physiologically reactive substance with which to block and/or remove toxic dicarbonyls and/or carbonyls from a living system.
It is an object therefore of the present invention to provide a method of removing toxic carbonyls and/or dicarbonyls, for example from a living body by administering a therapeutically effective dose of L- and/or D-arginine or an arginine-containing compound to a living body, the arginine thereby chemically reacting with the carbonyl group and preventing its reaction with native tissues.
It is a further object of the present invention to provide a method of preventing, alleviating or reducing complications associated with toxic carbonyls and/or dicarbonyls forming adducts and/or cross-links with native tissues by the therapeutic administration of L- and/or D-arginine or arginine-containing compounds to prevent such complex formation.
It is a further object of the present invention to provide a method of treating, for example, complications arising from cross-linking and/or adduct formation caused by toxic carbonyls and/or dicarbonyl-containing sugar metabolites such as methylglyoxal in diseases such as diabetes mellitus, by administering a therapeutically effective dose of L- and/or D-arginine, or substituted or modified arginine, or arginine-containing compounds to a living body.
Further, it is an object of the present invention to utilize non-naturally occurring and non-physiologically reactive D-arginine as the toxic dicarbonyl and/or carbonyl blocking and/or reacting agent of the present invention.